A Disulfide Bridge Allows for Site‐Selective Binding in Liver Bile Acid Binding Protein Thereby Stabilising the Orientation of Key Amino Acid Side Chains

Abstract

The presence of a disulfide bridge in liver bile acid binding protein (L-BABP/S-S) allows for site-selective binding of two bile acids, glycochenodeoxycholic (GCDA) and glycocholic acid (GCA), differing only in the presence of a hydroxyl group. The protein form devoid of the disulfide bridge (L-BABP) binds both bile salts without discriminating ability. We investigate the determinants of the molecular recognition process in the formation of the heterotypic L-BABP/S-S complex with GCDA [corrected] and GCA [corrected] located in the superficial and inner protein sites, respectively. The comparison of the NMR spectroscopy structure of heterotypic holo L-BABP/S-S, the first reported for this protein family, with that of the homotypic L-BABP complex demonstrates that the introduction of a S-S link between adjacent strands changes the conformation of three key residues, which function as hot-spot mediators of molecular discrimination. The favoured χ(1) rotameric states (t, g(+) and g(-) for E99, Q100 and E109 residues, respectively) allow the onset of an extended intramolecular hydrogen-bond network and the consequent stabilisation of the side-chain orientation of a buried histidine, which is capable of anchoring a specific ligand.